Effects of cold anaesthesia on the defensive behaviour of honeybees

Cold anaesthesia is commonly used to temporarily immobilise honeybees and other insects for a wide range of experimental investigations. Although cold-narcosis is widely regarded as non-invasive and non-damaging for neural structures, it has been shown that it can influence the bees’ subsequent behaviour in various ways. While we do have some understanding of the effect of cold treatment on handling stress, memory, locomotion and foraging in bees, it remains to be tested whether their defensive behaviour is affected. We investigated the response of foragers to a leg pinch applied before, during or after cold-anaesthesia. As expected, bees pinched at the hive entrance all responded defensively, while anaesthetised bees showed no response. When the pinch was applied after the bees recovered from cold-narcosis, all individuals were defensive, suggesting that previous cooling had not impaired nociception. We then subjected two groups of bees to a continuous pinch, commencing either before or during cold anaesthesia. Interestingly, most of these bees did not show defensive behaviour once awake, even though the clip was still attached to their leg. Chilling only lead to a diminished defensive response if the nociceptive stimulus was continuous rather than momentary, which suggests that it is not the chilling on its own, but the duration of the pinch in combination with cold narcosis that is critical. Our findings highlight the subtle ways in which treatment of insects during narcosis can affect their subsequent behaviour, and point to new ways in which narcosis can interact with sensory perception and adaptation.